Cellulose and cellulose derivatives have been used as chromatographic supports and as polymeric carriers. General chromatographic uses include analytical and preparative column chromatography, thin layer chromatography, ion exchange, gel chromatography and chelation and affinity sorbents. In addition, cellulose particles may be used as fillers and bulking agents in pharmaceuticals, cosmetics, and food products. Although natural abundance and availability, coupled with a variety of known derivatization schemes, make cellulose an attractive chromatographic support, it has generally suffered from several disadvantages. The most notable disadvantages are mechanical instability and poor flow characteristics.
Cellulose is a naturally occurring polymer made of linked glucose monomers. In the native state, adjacent polymeric glucose chains are extensively hydrogen bonded in some regions and less hydrogen bonded in others. The regions of relatively high hydrogen bonding are generally referred to as "microcrystalline regions" while the less hydrogen bonded regions are referred to as amorphous regions. For chromatographic applications, it is generally desirable to limit the amorphous regions, and to utilize cellulose having either a fibrous or a microgranular form. Such fibrous and microgranular materials are generally prepared by limited acid hydrolysis of bulk cellulose which results in the preferential loss of interchain amorphous regions and increases the microcrystalline regions. Both fibrous and microgranular cellulose compositions are generally referred to as microcrystalline cellulose.
Procedures typically used to prepare microcrystalline cellulose generally result in aggregated particles which require grinding and particle size separation to yield materials suitable for chromatographic purposes. Further, the individual microcrystalline cellulose particles are relatively irregularly shaped and fragile. These features adversely effect the use of these types of materials as chromatographic beds or columns because microcrystalline cellulose tends to easily clog and compact. In addition, these materials tend to break down and generate fines when subject to elevated pressure. These drawbacks can result in unacceptable flow characteristics and poor chromatographic separations.
The use of cellulose in the form of crosslinked beads or spherical particles may partially overcome the poor flow characteristics of microcrystalline cellulose chromatographic supports. When cellulose is made into beads using known procedures, however, the porosity of the cellulose particles and their wide range of sizes cause the beads to be subject to mechanical breakdown when used in packed beds or columns. Although, mechanical stability may be improved by the addition of cross linking agents, these crosslinking agents may increase the expense of the support, complicate the manufacturing processes and limit the general applicability of use of the support.
Further, when placed in aqueous solutions, porous cellulose particles typically swell significantly. Swelled, porous cellulose beads suffer from sensitivity to changing ionic strengths in eluting buffers and solvents. As a result, conventional, swellable cellulose supports must therefore be used within a specified narrow range of ionic strengths. If this specific range of ionic strengths is exceeded, the swelled cellulose particles compact or shrink which results in very poor flow characteristics and leads to either poor chromatographic separation or to no separation at all.
Several methods of preparing spherical cellulose particles are known. One method for preparing cellulose beads extrudes a viscose at high speed through a nozzle into a spinning acidic coagulating bath. Another method forms a dispersion in an organic solvent with a surfactant and then coagulates the suspension by pouring it into an acid solution. These procedures generate porous cellulose particles of variable and uncontrolled size distribution and suffer from the undesired formation of aggregates, agglomerates, and conglomerates of irregular and deformed shapes. These problems are believed due to the coagulation of the cellulose under changing hydrodynamic conditions. A third procedure thermally forms cellulose particles by heating an aqueous suspension of low molecular weight sodium cellulose xanthate in a stirred, relatively low viscosity, water-immiscible liquid. Although this procedure uses a relatively-constant, hydrodynamic environment for bead formation, the thermal decomposition of the sodium cellulose xanthate results in porous particles having a wide range of sizes.
There exists a need for a spherical, high density cellulose chromatographic support which has excellent mechanical stability, which may be used over a wide range of pH values and ionic strengths, and which may be readily prepared by a reproducible general method which provides uniformly sized and shaped particles.